Identification of tyrosine sulfation in extracellular leucine-rich repeat proteins using mass spectrometry
Identification of tyrosine sulfation in extracellular leucine-rich repeat proteins using mass spectrometry
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DOI：
10.1074/jbc.m308689200
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发表时间：
2004-01-02
影响因子：
4.8
通讯作者：
Heinegård, D
中科院分区：
文献类型：
--
作者：
Önnerfjord, P;Heathfield, TF;Heinegård, D
用质谱仪分析了胞外II类富含亮氨酸的重复蛋白/蛋白多糖中多种可变酪氨酸的硫化作用。酪氨酸上的磺基是不稳定的，在正常的质谱学条件下从多肽中释放出来。因此，必须考虑采用特殊方法来确定这一修改。通过使用在负离子和正离子模式下运行的质谱学研究的组合，可以确定酪氨酸的硫化。在阳性模式下，多肽通常表现为非硫酸盐化，而在阴性模式下，观察到硫酸盐和非硫酸盐化物种的混合物。不同酶释放的多肽的组合被用来获得硫酸盐基团位置的详细信息。在纤维调素(多达9个位点)、骨粘着蛋白(多达6个位点)和鲁米肯(2个位点)的N-末端区域观察到多种酪氨酸硫酸盐。骨粘着蛋白在其C末端附近含有两个额外的硫化酪氨酸残基。我们还发现了已发表的牛纤维调节蛋白序列中的一个错误，导致Thr(37)被Tyr(37)-Gly(38)取代，从而增加了它与人类同行的同源性。
Multiple and variable tyrosine sulfation in extracellular class II leucine-rich repeat proteins/proteoglycans were characterized by mass spectrometry. The sulfogroup on tyrosine is labile and is released from peptides under normal mass spectrometric conditions. Thus, special approaches must be considered in order to identify this modification. By using a combination of mass spectrometry studies operating in negative and positive ion mode, tyrosine sulfation could be identified. In positive mode, the peptides normally appeared non-sulfated, whereas in negative mode a mixture of sulfated and non-sulfated species was observed. A combination of peptides released by different proteinases was used to obtain details on the locations of sulfate groups. Multiple tyrosine sulfates were observed in the N-terminal region of fibromodulin ( up to 9 sites), osteoadherin ( up to 6 sites), and lumican ( 2 sites). Osteoadherin contains two additional sulfated tyrosine residues close to its C terminus. We also identified an error in the published sequence of bovine fibromodulin, resulting in the replacement of Thr(37) by Tyr(37)-Gly(38), thus increasing its homology with its human counterpart.